This Program Project application is a multi-disciplinary laboratory and clinical research program focused on the cellular mechanisms underlying post-ischemic myocardial inflammation, emphasizing the role of redox- sensitive events in vascular endothelial cells and myocytes which mediate gene transcription and neutrophil (PMN) accumulation. The specific objectives are to determine: (1) the mechanisms responsible for activation of the pro-inflammatory nuclear transcription factors NfkappaB and AP-1 following ischemia/reperfusion (I/R), including the role of pathways dependent on the small GP-binding protein Rac-1, (2) the effects of inhibiting NfkappaB activation on microvascular PMN trafficking, myocardial injury and cellular apoptosis following I/R, (3) the molecular interactions which occur between the oxygen radical and nitric oxide (NO) generation pathways, and their consequences vis-a-vis myocardial and vascular injury after I/R, (4) the transcriptional, post-transcriptional, and post-translational regulation of inducible nitric oxide synthase (NOS2) during I/R and the contribution of NOS2-derived NO to myocardial injury, (5) the redox mechanisms responsible for the hypoxic control of the transcriptional factor hypoxia-inducible factor-1 (HIF-1) and the consequences of HIF-1 expression in intact ischemia/reperfused hearts via-a-vis myocardial protection and post-ischemic inflammation. The Program Project has 4 projects and 3 cores (Viral, Pathology/Animal, and Administrative/Statistical). The techniques to be utilized include molecular cloning, construction of promoter-reporter gene constructs, gene transfer by recombinant adenoviral vectors, immunohistology, fluorescence confocal microscopy, in videomicroscopy, measurement of oxygen radicals and NO by electron paramagnetic resonance (EPR), and use of transgenic and knockout mice, cultured cells, perfused hearts and in vivo animal models. We anticipate that this proposed Program Project will yield fundamental information about the cellular mechanisms involved in the early stages of post-ischemic inflammation and will provide novel therapeutic approaches to reducing post-ischemic myocardial injury.